PKG serves as an integral component of second messenger signaling. In smooth muscle, the PKG isoforms Iα and Iβ integrate the nitric oxide (NO) and natriuretic peptide-mediated signal transduction pathways by modulating intracellular Ca2+, cell contractility, and—ultimately—blood flow (Hofmann et al., 2009; Francis et al., 1999; Pfeifer et al., 1999). Understanding the regulatory mechanism of the PKG holoenzyme is central to guiding pharmacological discoveries relevant to prevention and treatment of vascular diseases. Direct pharmaceutical targeting of PKG Iα however, has remained unsuccessful, largely due to the enzyme's complex, multi-domain architecture.